Alpha adrenergic receptors are plasma membrane receptors which are located in the peripheral and central nervous systems throughout the body. They are members of a diverse family of structurally related receptors which contain seven putative helical domains and transduce signals by coupling to guanine nucleotide binding proteins (G-proteins).
The alpha adrenergic receptor family of adrenergic receptors (AR) consists of two groups: alpha-1 and alpha-2. Of the alpha-2 group, there are three distinct subtypes denoted alpha-2A, alpha-2B and alpha-2C. The subtypes are derived from different genes, have different structures, unique distributions in the body, and specific pharmacologic properties. (Due to localization of the genes to human chromosomes 10, 2 and 4, the alpha-2A, alpha-2B, and alpha-2C receptors have sometimes been referred to as alpha-2C10, alpha-2C2 and alpha-2C4 receptors, respectively). Like other adrenergic receptors, the alpha-2 receptors are activated by endogenous agonists such as epinephrine (adrenaline) and norepinephrine (noradrenaline), and synthetic agonists, which promote coupling to G-proteins that in turn alter effectors such as enzymes or channels.
The alpha-2 receptors couple to the Gi and Go family of G-proteins. Alpha-2 receptors modulate a number of effector pathways in the cell: inhibition of adenylyl cyclase (decreases cAMP), stimulation of mitogen activated protein (MAP) kinase, stimulation of inositol phosphate accumulation, inhibition of voltage gated calcium channels and opening of potassium channels. (Limbird, L. E. (1988) FASEB J 2, 2686–2695, Luttrell, L. M., van Biesen, T., Hawes, B. E., Della Rocca, G. J., and Luttrell, D. K., and Lefkowitz, R. J. (1998) in Catecholamines: Bridging Basic Science with Clinical Medicine (Goldstein, D. S., Eisenhofer, G., and McCarty, R., eds pp. 466–470, Academic Press). The alpha-2 receptors are expressed on many cell-types in multiple organs in the body including those of the central and peripheral nervous systems.
Alpha-2BAR have a distinct pattern of expression within the brain, liver, lung, and kidney, and recent studies using gene knockouts in mice have shown that disruption of this receptor effects mouse viability, blood pressure responses to alpha-2-AR agonists, and the hypertensive response to salt loading. See Link, R. E., Desai, K., Hein, L., Stevens, M. E., Chruscinski, A., Bernstein, D., Barsh, G. S., and Kobilka, B. K. (1996) Science 273, 803–805; Makaritsis, K. P., Handy, D. E., Johns, C., Kobilka, B., Gavras, I., and Gavras, H. (1999) Hypertension 33, 14–17).
It is known that the alpha-2BAR undergoes short-term agonist promoted desensitization (Eason, M. G. and Liggett, S. B. (1992) J. Biol. Chem. 267, 25473–25479). This desensitization is due to phosphorylation of the receptor, which evokes a partial uncoupling of the receptor from functional interaction with Gi/Go (Jewell-Motz, E. A. and Liggett, S. B. (1995) Biochem 34, 11946–11953; Kurose, H. and Lefkowitz, R. J. (1994) J. Biol. Chem. 269, 10093–10099). Such phosphorylation appears to be due to G protein coupled receptor kinases (GRKs), a family of serine/threonine kinases which phosphorylate the agonist-occupied conformations of many G-protein coupled receptors (Pitcher, J. A., Freedman, N. J., and Lefkowitz, R. J. (1998) Annu Rev Biochem 67, 692). The phosphorylation process serves to finely regulate receptor function providing for rapid adaptation of the cell to its environment. Desensitization may also limit the therapeutic effectiveness of administered agonists. For the α2BAR, phosphorylation of serines/threonines in the third intracellular loop of the receptor is dependent on the presence of a stretch of acidic residues in the loop that appears to establish the milieu for GRK function (Jewell-Motz, E. A. and Liggett, S. B. (1995) Biochem 34, 11946–11953).
There has been a considerable research effort to clone and sequence the alpha-2AR. For example, the gene encoding the alpha-2A, alpha-2B, alpha-2C subtypes has been cloned and sequenced. (Kobilka et al. Science 238, 650–656 (1987); Regan et al., Lomasney et al. Proc. Nat. Acad. Sci. 87, 5094–5098 (1994)). These receptors have also been named as alpha-2C10, alpha-2C2 and alpha-2C4, according to their location on chromosomes 10, 4 and 2.
A polymorphism occurring in the gene encoding the alpha-2BAR has been previously reported. This polymorphism has been described as a deletion of three glutamic acid residues in a highly acidic stretch of amino acids in the third intracellular loop of the receptor. (Heinonen, P., Koulu, M., Pesonen, U., Karvonen, M. K., Rissanen, A., Laakso, M., Valve, R., Uusitupa, M., and Scheinin, M. (1999) J Clin Endocrinol Metab 84, 2429–2433; Baldwin, C. T., Schwartz, F., Baima, J., Burzstyn, M., DeStefano, A. L., Gavras, I., Handy, D. E., Joost, O., Martel, T., Manolis, A., Nicolaou, M., Bresnahan, M., Farrer, L., and Gavras, H. (1999) Am J Hypertens 12, 853–857). However, no pharmacologic studies have been carried out to determine if this polymorphism alters receptor function.
Given the importance of the alpha-2BAR in modulating a variety of physiological functions, there is a need in the art for improved methods to identify polymorphisms and to correlate the identity of these polymorphisms with signaling functions of alpha-2BAR. The present invention addresses these needs and more by providing polynucleotide and amino acid polymorphisms, molecules, and methods for detecting, genotyping and haplotyping the polymorphisms in the alpha-2BAR. The present invention is useful for determining an individual's risk for developing a disease, assist the clinician in diagnosing and prognosing the disease. The present invention also provides methods for selecting appropriate drug treatment based on the identity of such polymorphism.